Eps8 is a Convergence Point Integrating EGFR and Integrin Trafficking and Crosstalk

Crosstalk between adhesion and growth factor receptors plays a critical role in tissue morphogenesis and repair, and aberrations contribute substantially to neoplastic disease. However, the mechanisms by which adhesion and growth factor receptor signalling are integrated, spatially and temporally, are unclear.We used adhesion complex enrichment coupled with quantitative proteomic analysis to identify rapid changes to adhesion complex composition and signalling following growth factor stimulation. Bioinformatic network and ontological analyses revealed a substantial decrease in the abundance of adhesion regulatory proteins and co-ordinators of endocytosis within 5 minutes of EGF stimulation. Together these data suggested a mechanism of EGF-induced receptor endocytosis and adhesion complex turnover.Combinatorial interrogation of the networks allowed a global and dynamic view of adhesion and growth factor receptor crosstalk to be assembled. By interrogating network topology we identified Eps8 as a putative node integrating α5β1 integrin and EGFR functions. Importantly, EGF stimulation promoted internalisation of both α5β1 and EGFR. However, perturbation of Eps8 increased constitutive internalisation of α5β1 and EGFR; suggesting that Eps8 constrains α5β1 and EGFR endocytosis in the absence of EGF stimulation. Consistent with this, Eps8 regulated Rab5 activity and was required for maintenance of adhesion complex organisation and for EGF-dependent adhesion complex disassembly. Thus, by co-ordinating α5β1 and EGFR trafficking mechanisms, Eps8 is able to control adhesion receptor and growth factor receptor bioavailability and cellular contractility.We propose that during tissue morphogenesis and repair, Eps8 functions to spatially and temporally constrain endocytosis, and engagement, of α5β1 and EGFR in order to precisely co-ordinate adhesion disassembly, cytoskeletal dynamics and cell migration.Abbreviations: ECM: Extracellular matrixEGF: Epidermal growth factorEGFR: Epidermal growth factor receptorEps8: Epidermal growth factor receptor pathway substrate 8GFR: Growth factor receptorIAC: Integrin-associated complexMAPK: Mitogen activated protein kinaseMEC: Mammary epithelial cellMEF: Mouse embryonic fibroblastPKCα: Protein kinase CαTIF: T-antigen immortalised fibroblast